1. mar. biol. Ass.

India, 2002, 44 (1&2) : 85 - 96

Changes in biochemical and mineral composition during ovarian

maturation in the spiny lobster, Panulirus homarus (Linnaeus)
M. Vijayakumaran and E.V. Radhakrishnan
Central Marine Fisheries Research Institute, Cochin, India.

Abstract
Variations in biochemical and mineral composition of muscle, hepatopancreasand ovary were
studied during ovarian maturation in the spiny lobster, Panulirus homarus. Among the organic
reserves in the muscle, lipid alone showed a decline during maturation. In contrast, all the
organic reserves in the hepatopancreas declined in the mature and spent lobsters. The decrease in hepatopancreatic reserves amounted to 22.92% (lipid), 0.67% (protein) and 33.33%
(carbohydrate) of the total of these organic materials deposited in the mature ovary. The
decrease in energy reserves in hepatopancreas and the corresponding increase in the mature
ovary indicates that hepatopancreas is the main source of glycolipoproteins contributing to
the vitellogenic processes.
In the muscle, concentrations of Ca, Na, P, Mg, Cu, Zn, Mn, Co and Cd increased with attainment of maturity and declined after spawning. However, K was in maximum concentration
(17.56 mg/g dry wt.) in immature and Fe (176.96 mg/g dry wt.) and Cr (18.7 mg/g dry wt.) in
spent stages. Na, Ca and Mg concentrations in the hepatopancreas increased while K and P
decreased in mature and spent lobsters. With the exception of Co, all the trace elements in the
hepatopancreas increased with maturation; Cu (3507.33mg/g dry wt.) and Cd (61.34mg/g
dry wt.) recording 10 and 17 fold increases. In the ovary, all the minerals and trace elements
increased significantly (p<0.01) up to the ripe stage and decreased after spawning.

Introduction
Changes in biochemical composition of
tissues and organs during ovarian maturation have been studied in many crustaceans. Hepatopancreas or midgut gland,
which is the site of protein synthesis, lipid
metabolism and the main storage organ,
plays a vital role in the supply of energy
reserves, especially lipid for maturation
process (Allan, 1972; Adiyodi, 1985;
Teshima et al., 1989). Mobilization of
protein from hepatopancreas during

maturation is not certain, while

hepatopancreatic sugars are used for
maturation process (Adiyodi, 1985). In
lobsters, knowledge of biochemical
changes during maturation relates only to
the presence of female specific proteins in
the haemolymph in spiny lobsters and
micropinacocytosis within the oocytes of
homarid and palinurid lobsters. These
studies suggest extraovarian synthesis of
lipovitellin or yolk protein in lobsters
(Byard and Aiken, 1984; Adiyodi, 1985).

M. Vijayakumaran & E. V.Radhakrishnan

86

Role of trace elements in maturation of

fish and other aquatic animals is not
known, but their requirements differ at
different stages of maturation (Berman
and Vitin, 1968).Very few studies reported
in fishes indicate that there is variable
accumulation of Na, K, Mg, Mn, Fe, Cu
and Zn in different tissues, especially ovary
during maturation.(Love, 1980).In aquatic
animals, minerals required for embryogenesis have to be provided in the egg or
are to be absorbed from the medium in
which it undergoes development. In the
eggs of the spiny lobster, Pnntilirus homarus
and the penaeid prawn, Penaeus indicus,
mineral requirements are met both by
parental contribution in the ova as well as
selective absorption from the medium
(Vijayakumaran, 1990). The paper evaluates biochemical and mineral changes
during ovarian maturation in the spiny
lobster, P. hornarus.
The authors are grateful to Prof.
(Dr.) Mohan Joseph Modayil, Director,
Central Marine Fisheries Research Institute, Kochi-14 and to Dr. P.V.
Ramachandran Nair, Joint Director (retired) CMFRI, Kochi-14 for the constant
encouragement.
Material and methods
The lobsters were collected from
Kovalam, 25 k m south of Chennai, from
catches of traditional non-mechanized
units. Live lobsters were sacrificed immediately after collection and tissues processed for chemical estimation. Mature P.
homarus with spermatophore deposition
were held in aquarium for one or two
days and sacrificed soon after deposition
of eggs to collect samples of spent lobsters.

Three maturity stages, Immature

(Stage 11, Mature (Stage 4) and spent
(Stage 5) described for P. homarus by Berry
(1971) were followed. The size of lobsters
varied from 90-120g (immature), 121-545g
(mature) and 225-520g (spent).
Tissue samples were dried at 60 C to
constant weight and homogenized and
dried again for 1-2 hours before storing in
airtight glass vials in desiccator. Aliquots
from these dried samples were taken for
chemical analysis. Whole protein was
estimated calorimetrically by modified
biuret
method
(Sumitra
and
Vijayakumaran, 1979), carbohydrate by
phenol-sulfuric acid method (Raymont et
al., 1964) and total lipids by methanolchloroform extraction (Bligh and Dyer,
1969).After initial wet ashing, the samples
were dry ashed at 525O C for two hours
and mineral analyses were done by the
method described for fish and other
marine products by Thompson (1969) in
a Perkin - Elmer Atomic Absorption Spectrophotometer (Model 2380).
The term concentration in the text
means percentage in wet tissues for proximate composition, percentage in dry
weight for biochemical and mg or pg/g
dry weight for minerals and trace elements. The quantity is expressed as g/
lOOg body weight for proximate composition and mg or mg/100g body weight
for minerals and trace elements.
~

Biochemical changes in muscle

Changes in concentration of water,

87

Changes in biochemical and mineral composition in Panulirus homarus (Linnaeus)

Biochemical changes in hepatopancreas

protein, lipid, carbohydrate and ash in

muscle are given in Table 1. While water
and protein concentrations were comparatively stable during maturation, lipid and
carbohydrate concentrations decreased
with maturity and the minimum values
were recorded in spent lobsters. When
expressed quantitatively in a unit weight
of lOOg body weight (Table 2), the quantity of lipids declined significantly (p <
0.05) at maturity and also between mature and spent lobsters while other parameters were more or less stable.

index reduced from 3.79 in

immature to 3.34 in mature and further
to 2.88 in spent lobsters (Table 1).Water
and ash concentrations increased marginally at maturity and markedly after
spawning. Protein and carbohydrate concentrations showed a significant upward
trend in mature lobsters and decreased
significantly in the spent ones. A reverse
trend, a marked decline at maturity and
increase after spawning was noticed in
lipid concentration. Quantitative expresL

Table 1. Biochemical changes in muscle, hepatopancreas and ovary during ovarian maturatiot~in P. homarus
(protein, lipid, carbohydrate and ash expressed as percentage ill dry weight).
Maturity
stage

Size (g)

Immature

102.202
8.77

% in wet

weight
28.062
0.36

Water

74.962
2.25

Carbohydrate

Ash

Protein

Lipid

81.402
1.30

10.302
1.70

1.612
0.71

7.292
0.66

6.322
0.36

(%)
Muscle

Ripe
Spent
Hepatopancreas
-

--

Immature

102.202
8.77

3.792
0.05

66.52-16.67

53.512
3.21

31.712
4.50

5.60-10.61

Spent

388.31~
126.59

2.88-10.51

72.28-14.12

47.04*
2.24

32.902
0.13

5 . 9 2 ~ 13.412
0.41
0.11

Ovary
Immature

102.202
8.77

0.202
0.08

82.11-11.86

50.992
1.23

34.88~
0.28

3.172
0.29

6.372
0.44

388.312
126.59

0.832
0.12

76.922
6.21

64.97-14.09

19.112
3.44

2.732
0.27

12.912
0.28

Ripe
Spent

M. Vijayakurnaran & E. V . Radhakrishnan

88

sion (Table 2) revealed a different trend.

Quantity of protein declined significantly
(p < 0.005) at maturity and more so after
spawning. Lipid showed a significant
reduction (p c 0.0005) at maturity and a
marginal increase in spent lobsters. Increase at maturity and reduction after
spawning were highly significant (p <
0.005) in the quantity of carbohydrates of
hepatopancreas. Total energy considerably declined at maturity and also after
spawning. Quantity of ash showed a
significant positive change at maturity and
remained so after spawning.
Biochemical changes in ovary
Gonadosomaticindex increased sharply
from 0.2 to 5.3 at maturity and declined
to 0.83 after spawning (Table 1). Maximum concentration of water was recorded
in immature ovary and indicated a declining trend in mature ovary and increased after spawning. Concentrations
of protein and carbohydrates increased
while that of lipid declined at maturity.
After spawning, lipid and carbohydrate
concentrations declined while protein
recorded a marginal and ash 3 times increase in concentration. Quantitatively,
all parameters showed high increment and
equally significant reduction at maturity
and spawning, respectively (Table 2).

below detectable limit (<O.OOlmg/g dry

wt.) in all stages. Quantitatively, all elements, except Fe, which declined, significantly increased at maturity (Fig.1. a, b &
c). Between mature and spent stages all
elements except Ca, P, Co, and Cr declined in quantity. The most striking
changes were recorded for Co, which was
not detectable at maturity and accumulated several fold in the spent stage and
for Mn, which increased 8-fold at maturity and declined 15-fold after spawning.
Mineral changes i n hepatopancreas
In hepatopancreas, concentration and
quantity of all minerals and trace elements,
except K, P, Co and Cr, increased significantly in mature stage and recorded sharp
reduction after spawning (Table 3 and
Fig. 2. a, b & c). As in muscle, Pb was
below detectable level in all stages and Co
was not detectable in mature stage. 10
and 17 fold increases were recorded for
Cu and Cd, respectively at maturity.
Maximum rates of change in quantity
during maturation and spawning were
recorded for Na, Cu, Cd and Co.

zoo
lwzP
0

rn

20

Mineral changes i n muscle

2
10

In general, mineral and trace elemental concentrations increased with maturity in the muscle and declined after
'pawning
3)' was at maximum
concentration in h m a t u r e stage and so
also Fe and Cr. In spent stage Pb was

"

I M 5

I M 5

MMaturity
5
MSt.q. S

I M 5

I M 5

I M

Fig. 1 (a, b & c). Quantitative changes in nrrric~rrr1.s

and trace elements in muscle during ovarian
maturation in P, homarus (values expressed in
100 g body weight)

Changes in biochemical and mineral composition in Panulirus homarus (Linnaeus)

89

Table 2. Quantitative changes in proximate composition of muscle, hepatopancreas and ovary during
ovarian maturation in P. homarus (values expressed in 1008 body weight). Values in parenthesis are "P"
values of tests of significance (student's "t") between mature and immature, and mature and spent.
Maturity
stage

Wet
weight
(g)

Dry
Weight
(g)

Water
(g)

Protein
(g)

Lipid
(g)

Carbo
Hydrate
(g)

Ash
(g)

Total
energy
(calmlated)

(KJ)
Muscle

Immature

28.062
0.36
(<0.05)

Ripe

28.832
1.01

Spent

29.57+
1.42
(4.005)
Hepatopancreas

Immature

Ripe
Spent

Immature

Spent

3.792
0.05
(<0.0005)
3.342
0.16
2.882
0.51
b0.05)

0.202
0.08

0.042
0.01

0.162
0.06

0.020~
0.005

(<0.0005)

(<0.0005)

(<0.0005)

(<0.0005)

0.83~

0.202

0.652

0.1302

0.0142
0.003

0.0010~
0.0003

(<0.0005) (<0.0005)

0.1102

0.005k

0.0032
0.001

1.04~
0.24

(<0.0005) (<0.0005)

0.0102

7.472

M. Vijayakumaran t3 E. V . Radhakrishnan

90

linear relationship was observed between

ovary weight and body weight and the
ovary weight increased with maturation
irrespective of the size of the lobster.

Mmlurity Stop.

Fig. 2 (a, b & c). Quantitative clzanges in rninerals

and trace elements in hepatopancreas during
ovarian tnaturation in P. lzornarus (values expressed in 100g body weight)

Mineral changes in ovary

All elements in immature and spent
ovaries could not be detected since extremely low quantities of materials were
available for analysis. In contrast to the
conditions in muscle and hepatopancreas,
concentration of all minerals analyzed
declined at maturity and further after
spawning (Table 3). With the exception of
Co, concentration of all trace elements
showed an increasing trend in the mature
ovary. In the spent ovary, Zn, Cu and Co
were lesser in concentration.

Concentration of water in the ovary

declined significantlyfrom 82.11 to 52.71%
at maturity due to progressive addition of
organic reserves in developing oocytes.
When organic matter so accumulated
were finally transferred to the spawned
ova, the concentration of water again
increased to 72.91% in the spent ovary.
Ovary and hepatopancreas are the
main lipid storage organs in crustacea
(Guary et al., 1974). The total lipids increase in the ovary during sexual maturation in P. homarus (present study),
P. polyphagus (George and Patel, 19561, in
the sand lobster, Thenus orientalis (Rahman,
1989) and in many other crustaceans. As
the lipid reserves are ultimately transferred
to the mature ova, spawning results in
heavy depletion of lipids in the ovary.
Main component of crustacean yolk,
however, is protein, the lipovitellin

Quantitatively, all minerals and trace

elements recorded highly significant ( p<
0.0005) increase in mature ovary and
declined in a similar way after spawning
(Fig. 3. a, b & c).
Discussion
Maturation of ovary in P. homarus was
accompanied by a marked increase in
ovary mass, total lipid, protein, carbohycrustsand ash. As in many
ceans (Pillay and Nair, 1971; Clarke, 1977,
Jackel et al., 1989: Teshima et al., 1989). no

Fig. 3 (a, b & c). Quantitative changes iin ~ninerals

and trace elements in ovary during maturation
in P. komarus (values expressed in 100g-body
weight)

Table 3. Changes in minerals and trace elements composition (ins or mg / g dry u~eight)in mlrscle, l~epatopar~crens
and ovary dltring ovarial1 mntltration

ill P. homarus ( N.D. denotes not detectable concentration).

Minerals/
elements
(conc.)
Minerals
(mg/g dry
weight)

Trace elements

(mg/g dry
weight)

Maturity stage

Immature

Muscle
Ripe

Spent

Hepatopancreas
Immature
Ripe

Spent

Immature

Ovary
Ripe

Spent

92

(Adiyodi, 1985) and in P. homarus also the

quantity of protein increases 78 fold (from
0'2
1'58g) in the
Ovary and
62.82% of the total dry matter. At the
same time, the lipid content increased only
52
from 0.014 to 0.75g. The highest
concentration of protein in the ovary
(64'97%) was recorded in the 'Pent 'Ondition? even though the
quantity
was
low'
for better
understanding of the
and
utilization of organic reserves during
maturation, values have to be quantified
rather than being
in Percentage.
Lipid in hepatopancreas of P. homarus
declined significantly in mature lobsters
with concomitant increase in the ovary.
This trend has earlier been reported in
many crustaceans leading to the conclusion that in crustaceans with well defined
hepatopancreas, energy reserves are
stored in it and are apparently used during vitellogenesis. Unlike lipid, maximum
reductions in quantities of protein and
carbohydrate were noticed between mature and 'pent stagesr which might p o s
sibly indicate the utilization of these reserves for increased metabolic activity
associated with spawning.
In many fishes, both muscle and liver
energy are depleted in the process of reproductive spending (Love, 1980), while
crustaceans are not generally believed to
mobilize energy from muscle for gonad
development. In this study, significant decrease was noticed in muscle lipid both at
maturity and after spawning which per-

M. Vijayakurnaran & E. V. Radhakrishnan

haps, would have been utilized for reproductive processes.

Accumulation or depletion of fresh
weight dry matter, organic matter, energy and ash in muscle, hepatopancreas
and ovary during maturation are summarized in Table
indicating the extent of
involvement of muscle and hepatopancress in ovarian maturation. As described
earlier, the role of muscle in the reproductive process in P. homarus appears to be
restricted to supplying small quantity of
lipid. In contrast, all organic and inorganic reserves in hepatopancreas declined
at maturity and still further after spawning. The most significant reduction
(42.50%) at maturity in hepatopancreas
was that of lipid, while marked decline
(40.62%)in protein quantity was noticed
at spawning. Total carbohydrates were
also reduced at maturity and at spawning, but energetically its contribution was
negligible. Likewise, 7.07KJ (21.42% of
total) of hepatopancreatic energy was
spent for maturation and another 5.6.KJ
(16.96%) of the total during spawning.
These results suggest the important role
of hepatopancreas in storage and mobilization of energy during ovarian
tion in P. homarus. But to what extent the
hepatopancreatic reserves contribute to
the total reproductive output have to be
evaluated in the right perspective. The
decline in quantities of lipid, protein and
carbohydrate in hepatopancreas amounts
only to 22.92%, 0.67% and 33.33%, respectively, of the total amount of these
organic materials deposited in the mature

93

Changes in biochemical and mineral composition in Panulirus homarus (Linnaeus)

Table 4. Summary of quantitative changes in organic and inorianic reserves in muscle, hepatopancreas and ovary
during maturation in Panulirus homarus (values expressed in 100 g body weight).
-

--

Quantitative changes
Parameters
Muscle
Wet weight (g)
Dry weight (g)
water (g)
Protein (g)
Lipid (g)
Carbohydrate(@
Ash (g)
Energy (KJ)
Na (mg)
K (mg)
Ca (mg)
P (mg)
Mg (mg)
Fc (pg)
Cu (pg)
Zn (pg)
Cd (pg)
Co (pg)
Mn (pg)
Cr (pg)

Immature to ripe
Hepa topancreas

4.77
+0.28

-0.45
-0.18

ovary. This important point is glaringly

omitted in most of the studies, which
describes mobilization of hepatopancreatic
reserves during maturation in many crustaceans. The bulk of organic reserves
deposited in the ovary during maturation
should, therefore, come through transformation of ingested food either directly from
the gut,
the h a e m o l ~ m ~ash in
the case of echinoderms (Giese, 1959) or
through the mediation of tissues like
hepatopancrease which is the most important site of protein synthesis in crustaceans
and O' Canner, 1983).
The importance of minerds and trace
elements in maturation process has not
been studied in detail in crustaceans. Most

Ovary
+5.11
+2.47

Muscle

Ripe to spent
Hepatopancreas

+0.74
M.17

-0.46
-0.29

Ovary
4.48
-2.31

of the investigations on uptake of elements

from water have been intended to know
osmotic ion regulation in varying salinities, toxicity to heavy metal concentration, nutritional requirements for growth
and bio-accumulation in soft edible tissues.

Trace elements are generally required

in minute amounts, with the possible
exception of Mg and Fe and are
in specific physiological activities (Hoar,
1975). In
homarus, Mg is present in
higher concentration in muscle, hepatopancreas and ovary justifying its classification among the major elements or minerals in this study. Concentrations of Cu,
Fe and Zn also were high, especially at

94

mature stage, while all other elements

analyzed were present in minute quantities. Pb was not even detectable ( <
0.001mg/g dry wt.) which might possibly be due to the exclusion of exoskeleton
in this study, as exoskeleton is reported to
sequester most of the Pb in crustacea to
be expelled at the time of moulting (Eisler,
1981).
The quantity of K is reduced at maturity in muscle and hepatopancreas, and P
significantly reduced in hepatopancreas.
Other minerals (Na, Ca and Mg) accumulated in muscle, hepatopancreas and ovary
during maturation. A similar trend in
depletion of K and P during maturation
was reported in the penaeid shrimp, P.
indicus (Vijayakumaran, 1990).

M. Vijayakumaran & E. V. Radhakrishnan

bly to prevent them from being toxic to

the animal. Crustaceans expel excess
metal through exuvia and to a limited
extent through egg production (Davis,
1978). In P. homarus, the loss in quantity
of trace elements in muscle and hepatopancreas could not be fully accounted for
in its deposition in the spawned egg
(Vijayakumaran, 1990). Probably, these
elements would have been utilized for
increased metabolic activity related to
spawning.
Notable reduction in Co in muscle and
hepatopancreas in P. homarus and P.
indicus at maturity and selective absorption and utilization during embryogenesis
in these two species (Vijayakumaran,
1990) point to the importance of Co during maturation and egg development. Fe,
Zn and Cu were the most important trace
elements, in terms of quantity, in the
mature ovary and also just spawned eggs
of P. homarus and P. indicus
(Vijayakumaran, 1990). These elements
are important components of various
enzymes, including cytochrome oxidase
and blood pigment and play a major role
in the embryonic metabolism of the egg.

Cu, Zn, Fe, Cd and Mn quantitatively

accumulated in muscle, hepatopancreas
and ovary at maturity. The accumulation
of Cu in hepatopancreas was so high
(794.91%) at maturity that the ash was
light blue in colour. However, maximum
increase (1328.68%) was recorded in the
quantity of Cd, which is generally considered as non-essential and toxic
(Fleischer et al., 1974). Maximum loss of
In the fish, Rutilus rutilus, Ilzinia (1968)
trace elements (> 80%) after spawning in
hepatopancreas was also recorded for Cu noted variable accumulation of Mn, Fe,
and Cd. The loss of trace elements due Cu and Zn in various tissues, especially
to spawning was high in hepatopancreas ovary, with maturation. After spawning,
compared to muscle indicating its role in the concentration reduced in the ovary as
maturation process. Hepatopancreas, in well as in other tissues like muscle, liver,
crustaceans, is reported to sequester and bones and scales. A similar trend of
accumulate many metals like Cu, Cd and depletion of inorganic reserves in tissues
Zn (Eisler, 1981, Bjerregard and Vislie, after spawning, now reported in P.
1968) in bound and inactive form, possi- homarus indicates the importance of min-

Changes in biochemical and mineral composition in Panulirus homarus (Linnaeus)

era1 accumulation during maturation and

calls for further studies to understand the
role of individual minerals and trace elements in the maturation process.

95

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Marine Biology. Russel, S.F. and Yonge, S.M.
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York

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